Determining the Subcellular Localization of Fluorescently Tagged Proteins Using Protoplasts Extracted from Transiently Transformed Nicotiana benthamiana Leaves.
Identifieur interne : 000187 ( Main/Exploration ); précédent : 000186; suivant : 000188Determining the Subcellular Localization of Fluorescently Tagged Proteins Using Protoplasts Extracted from Transiently Transformed Nicotiana benthamiana Leaves.
Auteurs : Vivien Rolland [Australie]Source :
- Methods in molecular biology (Clifton, N.J.) [ 1940-6029 ] ; 2018.
Descripteurs français
- KwdFr :
- Cytoplasme (métabolisme), Espace intracellulaire (métabolisme), Feuilles de plante (génétique), Feuilles de plante (métabolisme), Microscopie de fluorescence (MeSH), Phénotype (MeSH), Protéines de fusion recombinantes (génétique), Protéines de fusion recombinantes (métabolisme), Protéines végétales (génétique), Protéines végétales (métabolisme), Tabac (génétique), Tabac (métabolisme), Transport des protéines (MeSH).
- MESH :
English descriptors
- KwdEn :
- Cytoplasm (metabolism), Intracellular Space (metabolism), Microscopy, Fluorescence (MeSH), Phenotype (MeSH), Plant Leaves (genetics), Plant Leaves (metabolism), Plant Proteins (genetics), Plant Proteins (metabolism), Protein Transport (MeSH), Recombinant Fusion Proteins (genetics), Recombinant Fusion Proteins (metabolism), Tobacco (genetics), Tobacco (metabolism).
- MESH :
- chemical , genetics : Plant Proteins, Recombinant Fusion Proteins.
- genetics : Plant Leaves, Tobacco.
- metabolism : Cytoplasm, Intracellular Space, Plant Leaves, Plant Proteins, Recombinant Fusion Proteins, Tobacco.
- Microscopy, Fluorescence, Phenotype, Protein Transport.
Abstract
In plants, stable expression is arguably the method of choice to test transgene function but it is a slow and labor-intensive process. This bottleneck generally limits the number of transgenes that can be tested, and as such hinders construct optimization. In the face of this challenge, transient expression in Nicotiana benthamiana leaves has emerged as a powerful screening platform to test gene expression, as well as subcellular distribution and function of many proteins within a week. This system relies on the infiltration of Agrobacterium tumefaciens (Agrobacterium) carrying DNA of interest into the leaf air spaces of N. benthamiana plants. Agrobacterium rapidly transforms the plant cells and the leaves can be analyzed within a few days. Investigating the subcellular localization of a protein of interest often relies on its fusion to a fluorescent tag. While the amount of accumulation of such fusion proteins can often be gauged by observing the fluorescence of the tag at the whole-leaf level, subcellular protein distribution is best determined in protoplasts extracted from transformed leaves. Here I present a simple and effective method to transform N. benthamiana leaves with Agrobacterium and to prepare protoplasts from these leaves to characterize the subcellular localization of proteins of interest.
DOI: 10.1007/978-1-4939-7786-4_16
PubMed: 29978408
Affiliations:
Links toward previous steps (curation, corpus...)
Le document en format XML
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(genetics)</term><term>Tobacco (metabolism)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Cytoplasme (métabolisme)</term><term>Espace intracellulaire (métabolisme)</term><term>Feuilles de plante (génétique)</term><term>Feuilles de plante (métabolisme)</term><term>Microscopie de fluorescence (MeSH)</term><term>Phénotype (MeSH)</term><term>Protéines de fusion recombinantes (génétique)</term><term>Protéines de fusion recombinantes (métabolisme)</term><term>Protéines végétales (génétique)</term><term>Protéines végétales (métabolisme)</term><term>Tabac (génétique)</term><term>Tabac (métabolisme)</term><term>Transport des protéines (MeSH)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Plant Proteins</term><term>Recombinant Fusion Proteins</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Plant Leaves</term><term>Tobacco</term></keywords><keywords scheme="MESH" qualifier="génétique" 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xml:lang="en">In plants, stable expression is arguably the method of choice to test transgene function but it is a slow and labor-intensive process. This bottleneck generally limits the number of transgenes that can be tested, and as such hinders construct optimization. In the face of this challenge, transient expression in Nicotiana benthamiana leaves has emerged as a powerful screening platform to test gene expression, as well as subcellular distribution and function of many proteins within a week. This system relies on the infiltration of Agrobacterium tumefaciens (Agrobacterium) carrying DNA of interest into the leaf air spaces of N. benthamiana plants. Agrobacterium rapidly transforms the plant cells and the leaves can be analyzed within a few days. Investigating the subcellular localization of a protein of interest often relies on its fusion to a fluorescent tag. While the amount of accumulation of such fusion proteins can often be gauged by observing the fluorescence of the tag at the whole-leaf level, subcellular protein distribution is best determined in protoplasts extracted from transformed leaves. Here I present a simple and effective method to transform N. benthamiana leaves with Agrobacterium and to prepare protoplasts from these leaves to characterize the subcellular localization of proteins of interest.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">29978408</PMID><DateCompleted><Year>2019</Year><Month>02</Month><Day>11</Day></DateCompleted><DateRevised><Year>2019</Year><Month>02</Month><Day>15</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Electronic">1940-6029</ISSN><JournalIssue CitedMedium="Internet"><Volume>1770</Volume><PubDate><Year>2018</Year></PubDate></JournalIssue><Title>Methods in molecular biology (Clifton, N.J.)</Title><ISOAbbreviation>Methods Mol Biol</ISOAbbreviation></Journal><ArticleTitle>Determining the Subcellular Localization of Fluorescently Tagged Proteins Using Protoplasts Extracted from Transiently Transformed Nicotiana benthamiana Leaves.</ArticleTitle><Pagination><MedlinePgn>263-283</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.1007/978-1-4939-7786-4_16</ELocationID><Abstract><AbstractText>In plants, stable expression is arguably the method of choice to test transgene function but it is a slow and labor-intensive process. This bottleneck generally limits the number of transgenes that can be tested, and as such hinders construct optimization. In the face of this challenge, transient expression in Nicotiana benthamiana leaves has emerged as a powerful screening platform to test gene expression, as well as subcellular distribution and function of many proteins within a week. This system relies on the infiltration of Agrobacterium tumefaciens (Agrobacterium) carrying DNA of interest into the leaf air spaces of N. benthamiana plants. Agrobacterium rapidly transforms the plant cells and the leaves can be analyzed within a few days. Investigating the subcellular localization of a protein of interest often relies on its fusion to a fluorescent tag. While the amount of accumulation of such fusion proteins can often be gauged by observing the fluorescence of the tag at the whole-leaf level, subcellular protein distribution is best determined in protoplasts extracted from transformed leaves. Here I present a simple and effective method to transform N. benthamiana leaves with Agrobacterium and to prepare protoplasts from these leaves to characterize the subcellular localization of proteins of interest.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Rolland</LastName><ForeName>Vivien</ForeName><Initials>V</Initials><AffiliationInfo><Affiliation>Commonwealth Scientific and Industrial Research Organisation (CSIRO), Agriculture & Food, Canberra, 2601, ACT, Australia. vivien.rolland@csiro.au.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>Methods Mol Biol</MedlineTA><NlmUniqueID>9214969</NlmUniqueID><ISSNLinking>1064-3745</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D010940">Plant Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D011993">Recombinant Fusion Proteins</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D003593" MajorTopicYN="N">Cytoplasm</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D042541" MajorTopicYN="N">Intracellular Space</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D008856" MajorTopicYN="N">Microscopy, Fluorescence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D010641" MajorTopicYN="N">Phenotype</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D018515" MajorTopicYN="N">Plant Leaves</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010940" MajorTopicYN="N">Plant Proteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D021381" MajorTopicYN="N">Protein Transport</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D011993" MajorTopicYN="N">Recombinant Fusion Proteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D014026" MajorTopicYN="N">Tobacco</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading></MeshHeadingList><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="Y">Agrobacterium tumefaciens</Keyword><Keyword MajorTopicYN="Y">Agroinfiltration</Keyword><Keyword MajorTopicYN="Y">Fluorescent proteins</Keyword><Keyword MajorTopicYN="Y">GFP</Keyword><Keyword MajorTopicYN="Y">Nicotiana benthamiana</Keyword><Keyword MajorTopicYN="Y">Protein targeting</Keyword><Keyword MajorTopicYN="Y">Protoplasts</Keyword><Keyword MajorTopicYN="Y">Subcellular localization</Keyword><Keyword MajorTopicYN="Y">Transient expression</Keyword></KeywordList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="entrez"><Year>2018</Year><Month>7</Month><Day>7</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2018</Year><Month>7</Month><Day>7</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2019</Year><Month>2</Month><Day>12</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">29978408</ArticleId><ArticleId IdType="doi">10.1007/978-1-4939-7786-4_16</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list><country><li>Australie</li></country></list><tree><country name="Australie"><noRegion><name sortKey="Rolland, Vivien" sort="Rolland, Vivien" uniqKey="Rolland V" first="Vivien" last="Rolland">Vivien Rolland</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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